Figure 3.

Examination of DNA binding mutants of Hop2–Mnd1 in the D-loop reaction. (a) Pulldown assay making use of Affi-Gel 15 Agarose beads coupled to DMC1 or BSA was conducted to test for the interaction with WT or mutant Hop2–Mnd1 complexes. The supernatant (S) containing unbound proteins, the wash (W) and the eluate (E) fractions were analyzed by SDS-PAGE and Coomassie Blue staining. (b) Schematic representation of the D-loop reaction. (c) WT and mutant variants of the Hop2–Mnd1 complex (60 nM or 120 nM) were tested for their ability to promote D-loop formation with ATP as nucleotide cofactor. The mean values ± SD from three independent experiments were quantified and plotted. (d) WT and mutant variants of the Hop2–Mnd1 complex (60 nM or 120 nM) were tested for their ability to promote D-loop formation with AMP-PNP as nucleotide cofactor. The mean values ± SD from three independent experiments were quantified and plotted.